Amphibian oocytes are large cells, 1.3 mm in diameter that are suitable for microinjection. A substantial body of evidence indicates that oocyte maturation is associated with a decrease in the phosphorylation state of a specific phosphoprotein inhibitor that is phosphorylated by cAMP-dependent protein kinase. The possibility that protein phosphatase Inhibitor-1 (Inh-1) is a phosphoprotein inhibitor will be assessed by Western blotting of oocytes with an antibody specific for phospho inhibitor 1 as well as microinjection of 32 Pi-labeled Inh-1. An activity in egg extracts that promotes nuclear envelope breakdown, chromosome condensation, and spindle formation will be purified and its regulation by Ca2+ ion and protein phosphorylation assessed by gel electrophoresis and microinjection studies. The system will be extended to mammalian somatic nuclei and permeabilized cells. The pattern of staining of centromeric regions on in vitro condensed chromosomes will be assessed in immunofluorescence studies using sera from patients with CREST schlerodoma. These experiments will provide new insights into the molecular basis of regulation of vertebrate oocyte maturation and cell division.